1. Field of the Invention
This invention relates to a method of producing a gel sheet for electrophoresis. More particularly, this invention relates to a method of providing a slot for loading samples into the gel sheet for electrophoresis.
2. Description of the Prior Art
An electrophoresis widely utilized is used for separating or analyzing high-molecular materials capable of having a charge in a solution such as proteins, nucleic acids and their decomposition products in a sheet medium, suc as gel membrane or filter paper, containing a buffer solution based upon the difference of charge and molecular weight.
Particularly, in gene engineering field, an important technique is the determination of base sequence of an isotope-labeled nucleic acid by utilizing autoradiography. In the electrophoretic operation for this purpose, a base-specific reaction product (mixture) of an isotope-labeled DNA or DNA fragment is allowed to migrate in parallel in electric field. Electrophoresis of a series of base-specific reaction products are, in general, carried out in one electrophoretic sheet, and plural electrophoretic patterns are formed in parallel. The electrophoretic migration patterns obtained as autoradiograms are compared with each other, and the base sequence is determined.
For conducting electrophoresis using gel membrane consisting of starch, polyacrylamide or the like as the electrophoretic medium, heretofoe, the person to conduct electrophoresis has to prepare each time a gel membrane on a flat plate such as glass plate. Such preparation work was troublesome, and it was a great load for the person to conduct electrophoresis.
Recently, a sheet material for electrophoresis called gel sheet (Japanese Patent KOKAI 126237/1984) has been commercially available. The gel sheet 1, as shown in Figure 4, is composed of the gel membrane 2 for electrophoresis, a base film 3 and a cover film 4 disposed on the front and the rear of the gel membrane 2 respectively, and a pair of spacers 5, 5 disposed along both sides of the gel membrane 2. The base film 3 and the cover film 4 of non-rigid electrical insulating material such as polyester film, are usually in contact with the gel membrane 2. An end of the gel membrane 2 is indented across most of the width so as to form a slot 6 for loading samples between the base film 3 and the cover film 4. A series of rectangular wells 7, 7, . . . ,7 for placing sample solutions are closely formed at the bottom 8 of the slot 6. The upper edge of the cover film 4 is slightly cut off so that a sample solution is easily put into the well 7. End projection 9 partitioning the slot 6 into respective wells 7 has a width of more than 1 mm, therefore, a space of a similar width is formed between respective electrophoretic migration regions called lanes. In case that the comparison of respective electrophoretic migration patterns is necessary, such as in the case of DNA base sequence analysis where the comparison is carried out as to a series of fragments having four kinds of bases (A, G, C, T), such space should be omitted or as small as possible. Therefore, a method of using a plate member having a sawtooth-shaped portion has been developed for such requirement, as shown in FIG. 5. The member 10 is called shark's teeth comb, and inverse V-shaped projections 11, 11, . . . 11 are formed at regular intervals in the teeth portion. In this method, the bottom 8 of the slot 6 is made straight, and the shark's teeth comb 10 is inserted into the slot 6, till respective tips of the inverse V-shaped projections 11 come into contact with or partly inserted into the upper edge of the gel membrane 2. Then, sample solutions are injected into each triangular or trapezoidal well formed by the projections of the shark's teeth comb 10 and the bottom 8 of the slot 6.
The above gel sheet 1 for electrophoresis is assembled, for example, by the process of FIG. 3 disclosed in Japanese Patent KOKAI No. 203847/1985, using either the gel sheet of FIG. 4 or the gel sheet of FIG. 5. In this process, a web of the base film 3 is delivered from its wound roll, and spacers 5, 5 delivered from their rolls are stuck to both side portions of the web of the base film 3. A gel-forming liquid composition is poured in film-shape from a hopper 12 on the web of the base film 3 so as to fill the whole space formed by the upper face of the web and two spacers 5, 5 with the liquid. Then, the gel-forming liquid composition is cured by passing it through a curing chamber 13 to form the gel membrane 2 for electrophoresis. The gel membrane 2 is punched by a punching means 14, and the cut piece of gel membrane is removed to form the portion corresponding to the "upper" (meaning that it is attached the upper end of upright type electrophoresis apparatus) edge of the gel membrane 2 in thegel sheet 1. A web of the cover film 4 is delivered from its wound roll, and stuck on the web of the gel membrane 2 supported by the base film 3. Then, the web is cut by the cutter 15 at each prescribed position to complete the gel sheet 1. In the above process of prior art, the punching process is carried out, as shown in FIG. 6. In the drawing, a half-assembled web moves intermittently in the leftward direction, and at each stop time, the pressure-controled chamber 17 of the punching means 16 descends. The gel membrane 2 is punched by a punching blade (not illustrated) provided on the underside of the pressure-controled chamber 17, and the cut piece of gel membrane 18 is adsorbed by the suction through the suction pipe 19. The pressure-controled chamber 17 holding the cut piece 18 ascends, and the receiving pan 20 comes under the pressure-controled chamber 17. After the suction pipe 19 is closed, the air supply pipe 21 connected to the pressure-controled chamber 17 is opened. While, the suction pipe 22 connected to the receiving pan 20 is opened, then the cut piece 18 is adsorbed to the receiving pan 20. The receiving pan 20 returns to the original place, and the cut piece 18 can be discharged by pressurizing through an air supply pipe 23. Thus, the procedure of one cycle is completed, and the next cycle is repeated.
In the above method, however, the suction was not effected uniformly depending on the position in the cut piece 18, and a fragment of the cut piece often remained on the gel sheet 1 as shown in the part X of FIG. 6. Moreover, a part of the cut piece 18 was stuck to the pressure control chamber 17, and could not be transferred to the receiving pan 20, as shown in the part Y of FIG. 6.
When such a state once happened, the working efficiency was remarkably lowered, since the fragment should be removed from the gel membrane 2 or the pressure control chamber 17 by hand.